Technical Abstract:
Around 1998, reassortment events occurred in pigs in the U.S. which resulted in the transfer of avian and human influenza genes into "classical" H1N1 (cH1N1) swine influenza viruses (SIV) producing novel H3N2, H1N2, H3N1 and reassortant H1N1 (rH1N1) viruses. These novel H3N2, H1N2 and rH1N1 viruses are now widespread in the U.S. swine population. Typically, the polymerase complex of these novel viruses consists of 2 avian-like polymerase genes (PA, PB2) and 1 human-like polymerase gene (PB1). We hypothesize that the novel viral polymerase genes are the basis for better adaptation and increased virulence of reassortant SIVs in pigs. To test this hypothesis, the eight-plasmid reverse genetic (rg) system was used to generate the triple reassortant H3N2 [A/Swine/Texas/4199-2/98 (Tx/98), rgTx98] and a cH1N1 SIV [A/Swine/IA/15/30 (IA/30), rg1930]. The rgTx98 was utilized to rescue reassortant SIVs possessing swine-like PA and/or PB2 polymerase genes from the double reassortant H3N2 [A/Swine/North Carolina/35922/98 (NC/98)] virus. The rg1930 system was employed to rescue reassortant SIVs that have avian-like and/or human-like polymerase gene(s) from Tx/98 virus instead of swine-like polymerase genes. The growth characteristics of the rescued viruses in MDCK, PK-15 and ST cells were analyzed and compared to that of the parental viruses. A total of ten reassortant SIVs were rescued and formed different size plaques. The reassortant viruses showed different growth characteristics in MDCK, PK-15 and ST cells. The reassortant IA/30-PB1/PB2 and Tx/98/NC-PA viruses showed a growth advantage in the three cell lines tested which was conferred by the combination of avian-like PB2 and human-like PB1 genes in conjunction with either an avian- or swine-like PA gene. Overall, these results suggest acquiring avian-like and/or human-like polymerase gene(s) plays an important role for replication in porcine cells and most likely for the adaptation and virulence of reassortant SIVs in pigs.